Steroid hormones regulate cell function by binding to cytoplasmic receptor proteins. these interact with acceptor sites in target cell nyuclei. There they induce messenger RNAs for specific cellular proteins which are responsible for alterations in endocrine physiologyl. Progesterone has been difficult to study at the molecular level in mammalian system. We have therefore characterized the progesterone-sensitive chick oviduct over the past 14 years and have shown it to be a representative model of steroid hormone action. During the course of the present grant, the structure and function of the chicken oviduct progesterone receptor will be studied in detail. This hormone binding protein is now available in purified form in amounts suitable for detailed protein structural determinations. The protein's subunit structure, DNA binding to the ovalbumin and ovomucoid genes, hormone-binding kinetics and mechanism, and effects on endocrine target cells will be studied in vitro. Also, its modification by cyclic AMP-dependent protein kinases will be stuied, with the goal of determining effects on function in oviduct cells. Proteolytic fragaments of both subunits A and B are to be sequenced. The hormone-binding site is to be studied by covalent labeling of the site by photo-activation of a synthetic ligand. Similarities or differences of sequence and secondary structure between A and B will be determined, together with detailed analysis of contacts between the two subunits in the intact complex. DNA interactions of subunit A will be defined more precisely, with respect to effects on DNA structure. Receptors in oviduct cells will be localized within the ovalbumin gene cluster by nuclease digestion and DNA Hybridization experiments. A cell-free oviduct transcription system will be developed for tests of effects of purified receptors in vitro. This work should afford us an opportunity to define precisely the molecular basis of hormone-mediated gene expression, resulting in an advance in our understanding of endocrine physiology, reproductive biology and steroid hormone action.